Fibroblast-specific PRMT5 deficiency suppresses cardiac fibrosis and left ventricular dysfunction in male mice.

Protein arginine methyltransferase 5 (PRMT5) is a well-known epigenetic regulatory enzyme. However, the role of PRMT5-mediated arginine methylation in gene transcription related to cardiac fibrosis is unknown. Here we show that fibroblast-specific deletion of PRMT5 significantly reduces pressure overload-induced cardiac fibrosis and improves cardiac dysfunction in male mice. Both the PRMT5-selective inhibitor EPZ015666 and knockdown of PRMT5 suppress α-smooth muscle actin (α-SMA) expression induced by transforming growth factor-ß (TGF-ß) in cultured cardiac fibroblasts. TGF-ß stimulation promotes the recruitment of the PRMT5/Smad3 complex to the promoter site of α-SMA. It also increases PRMT5-mediated H3R2 symmetric dimethylation, and this increase is inhibited by Smad3 knockdown. TGF-ß stimulation increases H3K4 tri-methylation mediated by the WDR5/MLL1 methyltransferase complex, which recognizes H3R2 dimethylation. Finally, treatment with EPZ015666 significantly improves pressure overload-induced cardiac fibrosis and dysfunction. These findings suggest that PRMT5 regulates TGF-ß/Smad3-dependent fibrotic gene transcription, possibly through histone methylation crosstalk, and plays a critical role in cardiac fibrosis and dysfunction.

Sex-based differences in short- and longer-term diet-induced metabolic heart disease.

Sex-based differences in the development of obesity-induced cardiometabolic dysfunction are well documented, however, the specific mechanisms are not completely understood. Obesity has been linked to dysregulation of the epitranscriptome, but the role of N6-methyladenosine (m6A) RNA methylation has not been investigated in relation to the sex differences during obesity-induced cardiac dysfunction. In the current study, male and female C57BL/6J mice were subjected to short- and long-term high-fat/high-sucrose (HFHS) diet to induce obesogenic stress. Cardiac echocardiography showed males developed systolic and diastolic dysfunction after 4 mo of diet, but females maintained normal cardiac function despite both sexes being metabolically dysfunctional. Cardiac m6A machinery gene expression was differentially regulated by duration of HFHS diet in male, but not female mice, and left ventricular ejection fraction correlated with RNA machinery gene levels in a sex- and age-dependent manner. RNA-sequencing of cardiac transcriptome revealed that females, but not males may undergo protective cardiac remodeling early in the course of obesogenic stress. Taken together, our study demonstrates for the first time that cardiac RNA methylation machinery genes are regulated early during obesogenic stress in a sex-dependent manner and may play a role in the sex differences observed in cardiometabolic dysfunction.NEW & NOTEWORTHY Sex differences in obesity-associated cardiomyopathy are well documented but incompletely understood. We show for the first time that RNA methylation machinery genes may be regulated in response to obesogenic diet in a sex- and age-dependent manner and levels may correspond to cardiac systolic function. Our cardiac RNA-seq analysis suggests female, but not male mice may be protected from cardiac dysfunction by a protective cardiac remodeling response early during obesogenic stress.

Chronic kidney disease and left ventricular diastolic dysfunction (CKD-LVDD) alter cardiac expression of mitochondria-related genes in swine.

Cardiovascular disease and heart failure doubles in patients with chronic kidney disease (CKD), but the underlying mechanisms remain obscure. Mitochondria are central to maintaining cellular respiration and modulating cardiomyocyte function. We took advantage of our novel swine model of CKD and left ventricular diastolic dysfunction (CKD-LVDD) to investigate the expression of mitochondria-related genes and potential mechanisms regulating their expression. CKD-LVDD and normal control pigs (n=6/group, 3 males/3 females) were studied for 14 weeks. Renal and cardiac hemodynamics were quantified by multidetector-CT, echocardiography, and pressure-volume loop studies, respectively. Mitochondrial morphology (electron microscopy) and function (Oroboros) were assessed ex vivo. In randomly selected pigs (n=3/group), cardiac mRNA-, MeDIP-, and miRNA-sequencing (seq) were performed to identify mitochondria-related genes and study their pre- and post -transcriptional regulation. CKD-LVDD exhibited cardiac mitochondrial structural abnormalities and elevated mitochondrial H2O2 emission but preserved mitochondrial function. Cardiac mRNA-seq identified 862 mitochondria-related genes, of which 69 were upregulated and 33 downregulated (fold-change ≥2, false discovery rate≤0.05). Functional analysis showed that upregulated genes were primarily implicated in processes associated with oxidative stress, whereas those downregulated mainly participated in respiration and ATP synthesis. Integrated mRNA/miRNA/MeDIP-seq analysis showed that upregulated genes were modulated predominantly by miRNAs, whereas those downregulated were by miRNA and epigenetic mechanisms. CKD-LVDD alters cardiac expression of mitochondria-related genes, associated with mitochondrial structural damage but preserved respiratory function, possibly reflecting intrinsic compensatory mechanisms. Our findings may guide the development of early interventions at stages of cardiac dysfunction in which mitochondrial injury could be prevented, and the development of LVDD ameliorated.

Thrombosis in the Coronary Microvasculature Impairs Cardiac Relaxation and Induces Diastolic Dysfunction.

BACKGROUND: Heart failure with preserved ejection fraction is proposed to be caused by endothelial dysfunction in cardiac microvessels. Our goal was to identify molecular and cellular mechanisms underlying the development of cardiac microvessel disease and diastolic dysfunction in the setting of type 2 diabetes. METHODS: We used Leprdb/db (leptin receptor-deficient) female mice as a model of type 2 diabetes and heart failure with preserved ejection fraction and identified Hhipl1 (hedgehog interacting protein-like 1), which encodes for a decoy receptor for HH (hedgehog) ligands as a gene upregulated in the cardiac vascular fraction of diseased mice. RESULTS: We then used Dhh (desert HH)-deficient mice to investigate the functional consequences of impaired HH signaling in the adult heart. We found that Dhh-deficient mice displayed increased end-diastolic pressure while left ventricular ejection fraction was comparable to that of control mice. This phenotype was associated with a reduced exercise tolerance in the treadmill test, suggesting that Dhh-deficient mice do present heart failure. At molecular and cellular levels, impaired cardiac relaxation in DhhECKO mice was associated with a significantly decreased PLN (phospholamban) phosphorylation on Thr17 (threonine 17) and an alteration of sarcomeric shortening ex vivo. Besides, as expected, Dhh-deficient mice exhibited phenotypic changes in their cardiac microvessels including a prominent prothrombotic phenotype. Importantly, aspirin therapy prevented the occurrence of both diastolic dysfunction and exercise intolerance in these mice. To confirm the critical role of thrombosis in the pathophysiology of diastolic dysfunction, we verified Leprdb/db also displays increased cardiac microvessel thrombosis. Moreover, consistently, with Dhh-deficient mice, we found that aspirin treatment decreased end-diastolic pressure and improved exercise tolerance in Leprdb/db mice. CONCLUSIONS: Altogether, these results demonstrate that microvessel thrombosis may participate in the pathophysiology of heart failure with preserved ejection fraction.

Human leukocyte antigen-DQ risk heterodimeric haplotypes of left ventricular dysfunction in cardiac sarcoidosis: an autoimmune view of its role.

Cardiac sarcoidosis (CS) is the scarring of heart muscles by autoimmunity, leading to heart abnormalities and patients with sarcoidosis with cardiac involvements have poor prognoses. Due to the small number of patients, it is difficult to stratify all patients of CS by human leukocyte antigen (HLA) analysis. We focused on the structure of antigen-recognizing pockets in heterodimeric HLA-class II, in addition to DNA sequences, and extracted high-affinity combinations of antigenic epitopes from candidate autoantigen proteins and HLA. Four HLA heterodimer-haplotypes (DQA1*05:03/05:05/05:06/05:08-DQB1*03:01) were identified in 10 of 68 cases. Nine of the 10 patients had low left ventricular ejection fraction (< 50%). Fourteen amino-acid sequences constituting four HLA anchor pockets encoded by the HLA haplotypes were all common, suggesting DQA1*05:0X-DQB1*03:01 exhibit one group of heterodimeric haplotypes. The heterodimeric haplotypes recognized eight epitopes from different proteins. Assuming that autoimmune mechanisms might be activated by molecular mimicry, we searched for bacterial species having peptide sequences homologous to the eight epitopes. Within the peptide epitopes form the SLC25A4 and DSG2, high-homology sequences were found in Cutibacterium acnes and Mycobacterium tuberculosis, respectively. In this study, we detected the risk heterodimeric haplotypes of ventricular dysfunction in CS by searching for high-affinity HLA-class II and antigenic epitopes from candidate cardiac proteins.

Insulin-like growth factor binding protein 2: a core biomarker of left ventricular dysfunction in dilated cardiomyopathy.

BACKGROUND: RNA modifications, especially N6-methyladenosine, N1-methyladenosine and 5-methylcytosine, play an important role in the progression of cardiovascular disease. However, its regulatory function in dilated cardiomyopathy (DCM) remains to be undefined. METHODS: In the study, key RNA modification regulators (RMRs) were screened by three machine learning models. Subsequently, a risk prediction model for DCM was developed and validated based on these important genes, and the diagnostic efficiency of these genes was assessed. Meanwhile, the relevance of these genes to clinical traits was explored. In both animal models and human subjects, the gene with the strongest connection was confirmed. The expression patterns of important genes were investigated using single-cell analysis. RESULTS: A total of 4 key RMRs were identified. The risk prediction models were constructed basing on these genes which showed a good accuracy and sensitivity in both the training and test set. Correlation analysis showed that insulin-like growth factor binding protein 2 (IGFBP2) had the highest correlation with left ventricular ejection fraction (LVEF) (R = -0.49, P = 0.00039). Further validation expression level of IGFBP2 indicated that this gene was significantly upregulated in DCM animal models and patients, and correlation analysis validation showed a significant negative correlation between IGFBP2 and LVEF (R = -0.87; P = 6*10-5). Single-cell analysis revealed that this gene was mainly expressed in endothelial cells. CONCLUSION: In conclusion, IGFBP2 is an important biomarker of left ventricular dysfunction in DCM. Future clinical applications could possibly use it as a possible therapeutic target.

Clonal Hematopoiesis in Clinical and Experimental Heart Failure With Preserved Ejection Fraction.

BACKGROUND: Clonal hematopoiesis (CH), which results from an array of nonmalignant driver gene mutations, can lead to altered immune cell function and chronic disease, and has been associated with worse outcomes in patients with heart failure (HF) with reduced ejection fraction. However, the role of CH in the prognosis of HF with preserved ejection fraction (HFpEF) has been understudied. This study aimed to characterize CH in patients with HFpEF and elucidate its causal role in a murine model. METHODS: Using a panel of 20 candidate CH driver genes and a variant allele fraction cutoff of 0.5%, ultradeep error-corrected sequencing identified CH in a cohort of 81 patients with HFpEF (mean age, 71±6 years; ejection fraction, 63±5%) and 36 controls without a diagnosis of HFpEF (mean age, 74±7 years; ejection fraction, 61.5±8%). CH was also evaluated in a replication cohort of 59 individuals with HFpEF. RESULTS: Compared with controls, there was an enrichment of TET2-mediated CH in the HFpEF patient cohort (12% versus 0%, respectively; P=0.02). In the HFpEF cohort, patients with CH exhibited exacerbated diastolic dysfunction in terms of E/e' (14.9 versus 11.7, respectively; P=0.0096) and E/A (1.69 versus 0.89, respectively; P=0.0206) compared with those without CH. The association of CH with exacerbated diastolic dysfunction was corroborated in a validation cohort of individuals with HFpEF. In accordance, patients with HFpEF, an age ≥70 years, and CH exhibited worse prognosis in terms of 5-year cardiovascular-related hospitalization rate (hazard ratio, 5.06; P=0.042) compared with patients with HFpEF and an age ≥70 years without CH. To investigate the causal role of CH in HFpEF, nonconditioned mice underwent adoptive transfer with Tet2-wild-type or Tet2-deficient bone marrow and were subsequently subjected to a high-fat diet/L-NAME (Nω-nitro-l-arginine methyl ester) combination treatment to induce features of HFpEF. This model of Tet2-CH exacerbated cardiac hypertrophy by heart weight/tibia length and cardiomyocyte size, diastolic dysfunction by E/e' and left ventricular end-diastolic pressure, and cardiac fibrosis compared with the Tet2-wild-type condition. CONCLUSIONS: CH is associated with worse heart function and prognosis in patients with HFpEF, and a murine experimental model of Tet2-mediated CH displays greater features of HFpEF.

Evaluation of left ventricular dysfunction by three-dimensional speckle-tracking echocardiography and bioinformatics analysis of circulating exosomal miRNA in obese patients.

BACKGROUND: Obesity is an independent risk factor for cardiovascular disease and affects the human population. This study aimed to evaluate left ventricular (LV) dysfunction in obese patients with three-dimensional speckle-tracking echocardiography (3D-STE) and investigate the possible related mechanisms at the exosomal miRNA level. METHODS: In total, 43 participants (16 obese patients and 27 healthy volunteers) were enrolled. All subjects underwent full conventional echocardiography as well as 3D-STE. Characterization and high-throughput sequencing for the isolated circulating exosomes and the differentially expressed miRNAs (DEMs) were screened for target gene prediction and enrichment analysis. RESULTS: Obese patients had significantly lower global longitudinal strain (GLS) (-20.80%±3.10% vs. -14.77%±2.05%, P < 0.001), global circumferential strain (GCS) (-31.63%±3.89% vs. -25.35%±5.66%, P = 0.001), global radial strain (GRS) (43.21%±4.89% vs. 33.38%±3.47%, P < 0.001), and indexed LV end-diastolic volume (LVEDV) [38.07mL/m2 (27.82mL/m2-9.57mL/m2) vs. 24.79mL/m2 (21.97mL/m2-30.73mL/m2), P = 0.002] than healthy controls. GLS (ρ = 0.610, P < 0.001), GCS (ρ = 0.424, P = 0.005), and GRS (ρ = -0.656, P < 0.001) indicated a moderate relationship with body mass index (BMI). In obese patients, 33 exosomal miRNAs were up-regulated and 26 exosomal miRNAs were down-regulated when compared to healthy controls (P < 0.05). These DEMs possibly contribute to obesity-associated LV dysfunction through the PI3K-Akt signaling pathway. Important miRNAs, including miR-101-3p, miR-140-3p, and miR-99a-5p, have clinical utility in predicting early obesity-related myocardial injury. CONCLUSIONS: The global strain obtained from 3D-STE can sensitively detect the decrease in LV myocardial function in obese patients. Key miRNAs and pathways provide a new theoretical basis and targets of action for studying obesity-induced LV dysfunction. TRIAL REGISTRATION: In accordance with the World Health Organization (WHO) definition of a clinical trial, this study does not include human health-related interventions. This study was carried out at the General Hospital of Ningxia Medical University after obtaining institutional ethical approval (KYLL-2022-0556) and written informed consent from all participants.

Risks of Ventricular Arrhythmia and Heart Failure in Carriers of RBM20 Variants.

BACKGROUND: Variants in RBM20 are reported in 2% to 6% of familial cases of dilated cardiomyopathy and may be associated with fatal ventricular arrhythmia and rapid heart failure progression. We sought to determine the risk of adverse events in RBM20 variant carriers and the impact of sex on outcomes. METHODS: Consecutive probands and relatives carrying RBM20 variants were retrospectively recruited from 12 cardiomyopathy units. The primary end point was a composite of malignant ventricular arrhythmia (MVA) and end-stage heart failure (ESHF). MVA and ESHF end points were also analyzed separately and men and women compared. Left ventricular ejection fraction (LVEF) contemporary to MVA was examined. RBM20 variant carriers with left ventricular systolic dysfunction (RBM20LVSD) were compared with variant-elusive patients with idiopathic left ventricular systolic dysfunction. RESULTS: Longitudinal follow-up data were available for 143 RBM20 variant carriers (71 men; median age, 35.5 years); 7 of 143 had an MVA event at baseline. Thirty of 136 without baseline MVA (22.0%) reached the primary end point, and 16 of 136 (11.8%) had new MVA with no significant difference between men and women (log-rank P=0.07 and P=0.98, respectively). Twenty of 143 (14.0%) developed ESHF (17 men and 3 women; log-rank P<0.001). Four of 10 variant carriers with available LVEF contemporary to MVA had an LVEF >35%. At 5 years, 15 of 67 (22.4%) RBM20LVSD versus 7 of 197 (3.6%) patients with idiopathic left ventricular systolic dysfunction had reached the primary end point (log-rank P<0.001). RBM20 variant carriage conferred a 6.0-fold increase in risk of the primary end point. CONCLUSIONS: RBM20 variants are associated with a high risk of MVA and ESHF compared with idiopathic left ventricular systolic dysfunction. The risk of MVA in male and female RBM20 variant carriers is similar, but male sex is strongly associated with ESHF.

Torsional and strain dysfunction precede overt heart failure in a mouse model of dilated cardiomyopathy pathogenesis.

Detailed assessments of whole heart mechanics are crucial for understanding the consequences of sarcomere perturbations that lead to cardiomyopathy in mice. Echocardiography offers an accessible and cost-effective method of obtaining metrics of cardiac function, but the most routine imaging and analysis protocols might not identify subtle mechanical deficiencies. This study aims to use advanced echocardiography imaging and analysis techniques to identify previously unappreciated mechanical deficiencies in a mouse model of dilated cardiomyopathy (DCM) before the onset of overt systolic heart failure (HF). Mice lacking muscle LIM protein expression (MLP-/-) were used to model DCM-linked HF pathogenesis. Left ventricular (LV) function of MLP-/- and wild-type (WT) controls were studied at 3, 6, and 10 wk of age using conventional and four-dimensional (4-D) echocardiography, followed by speckle-tracking analysis to assess torsional and strain mechanics. Mice were also studied with RNA-seq. Although 3-wk-old MLP-/- mice showed normal LV ejection fraction (LVEF), these mice displayed abnormal torsional and strain mechanics alongside reduced ß-adrenergic reserve. Transcriptome analysis showed that these defects preceded most molecular markers of HF. However, these markers became upregulated as MLP-/- mice aged and developed overt systolic dysfunction. These findings indicate that subtle deficiencies in LV mechanics, undetected by LVEF and conventional molecular markers, may act as pathogenic stimuli in DCM-linked HF. Using these analyses in future studies will further help connect in vitro measurements of the sarcomere function to whole heart function.NEW & NOTEWORTHY A detailed study of how perturbations to sarcomere proteins impact whole heart mechanics in mouse models is a major yet challenging step in furthering our understanding of cardiovascular pathophysiology. This study uses advanced echocardiographic imaging and analysis techniques to reveal previously unappreciated subclinical whole heart mechanical defects in a mouse model of cardiomyopathy. In doing so, it offers an accessible set of measurements for future studies to use when connecting sarcomere and whole heart function.

Layer myocardial strain is the most heritable echocardiographic trait.

AIMS: Myocardial deformation assessed by strain analysis represents a significant advancement in our assessment of cardiac mechanics. However, whether this variable is genetically heritable or whether all/most of its variability is related to environmental factors is currently unknown. We sought to determine the heritability of echocardiographically determined cardiac mechanics indices in a population setting. METHODS AND RESULTS: A total of 1357 initially healthy subjects (women 51.6%; 48.2 ± 14.1 years) were included in this study from 20-year follow-up after the fourth visit of the longitudinal familial STANISLAS cohort (Lorraine, France). Data were acquired using state-of-the-art cardiac ultrasound equipment, using acquisition and measurement protocols recommended by the EACVI (European Association of Cardiovascular Imaging)/ASE (American Society of Echocardiography)/Industry Task Force. Layer-specific global longitudinal strain (GLS) and global circumferential strain (full-wall, subendocardial, and subepicardial) and conventional structural and functional cardiac parameters and their potential heritability were assessed using restricted maximum likelihood analysis, with genetic relatedness matrix calculated from genome-wide association data. Indices of longitudinal/circumferential myocardial function and left ventricular (LV) ejection fraction had low heritability (ranging from 10% to 20%). Diastolic and standard LV function parameters had moderate heritability (ranging from 20% to 30%) except for end-systolic and end-diastolic volumes (30% and 45%, respectively). In contrast, global longitudinal subendocardial strain (GLSEndo)/global longitudinal subepicardial strain (GLSEpi) ratio had a high level of heritability (65%). Except for GLSEndo/GLSEpi ratio, a large percentage of variance remained unexplained (>50%). CONCLUSIONS: In our population cohort, GLSEndo/GLSEpi ratio had a high level of heritability, whereas other classical and mechanical LV function parameters did not. Given the increasing recognition of GLSEndo/GLSEpi ratio as an early/sensitive imaging biomarker of systolic dysfunction, our results suggest the possible existence of individual genetic predispositions to myocardial decline.

LncRNA HIF1A-AS2: a potential biomarker for early diagnosis of acute myocardial infarction and predictor of left ventricular dysfunction.

BACKGROUND: Rapid diagnosis of acute myocardial infarction (AMI) is the subject of many clinical studies as it enables an effective therapy, preventing adverse progression of AMI and increasing survival rates. Recent studies have revealed that specific blood-based long non-coding RNAs (lncRNAs) are deregulated in patients with AMI and serve as promising diagnostic and prognostic tools. The current study aimed to determine the potential role of a hypoxia-responsive lncRNA, hypoxia-inducible factor 1A antisense RNA 2 (HIF1A-AS2), as a biomarker for early diagnosis and predictor of left ventricular dysfunction (LVD). METHODS: This study was carried out on 48 patients with AMI and 50 age-and sex-matched controls. The relative quantification of HIF1A-AS2 expression was done using reverse transcription real-time polymerase chain reaction. RESULTS: Compared to the control group, HIF1A-AS2 were significantly higher in MI patients (P < 0.001). Interestingly, patients presenting within 3 h of chest pain onset had elevated levels of HIF1A-AS2 as compared to patients with late presentation. The ROC curve was constructed to assess HIF1A-AS2 as an early marker. It demonstrated higher sensitivity (94%) and specificity (86%). Moreover, the multivariate regression analysis revealed that HIF1A-AS2 was significantly associated with LVD in the patient group after 6 months follow up (p = 0.018). CONCLUSION: Our study suggests that HIF1A-AS2 may be a potential early diagnostic biomarker of AMI with high sensitivity. In addition, it might have a promising role as a predictor of left ventricular dysfunction.

Intrinsic Atrial Myopathy Precedes Left Ventricular Dysfunction and Predicts Atrial Fibrillation in Lamin A/C Cardiomyopathy.

BACKGROUND: In Lamin A/C (LMNA) cardiomyopathy, atrial fibrillation (AF) commonly occurs before dilated cardiomyopathy, and the ability to predict its incidence is limited. We hypothesized that left atrial (LA) echocardiographic phenotyping can identify atrial myopathy and harbingers of AF. METHODS: Echocardiograms from patients with pathogenic or likely pathogenic variants in LMNA (n=77) with and without reduced left ventricular ejection fraction (LVEF, <50%) were compared to healthy individuals (n=70) and patients with Titin truncating variant cardiomyopathy (TTNtv) (n=35) with similar LVEF, sex, and age distributions. Echocardiographic analysis, blinded to genotype, included strain and volumetric measures of left ventricular and atrial function. The primary outcome was incident AF. RESULTS: At baseline, 43% of the patients with pathogenic or likely pathogenic LMNA variants had a history of AF, including 26% of those with LVEF ≥50%. Compared with healthy subjects, the patients with pathogenic or likely pathogenic LMNA variants and LVEF ≥50% had reduced LA contractile strain (LMNA, 11.8±6.1% versus control, 15.0±4.2%; P=0.003). Compared to LVEF-matched (TTNtv) patients, the patients with pathogenic or likely pathogenic LMNA variants and LVEF <50% displayed no difference in LA size, but a worse LA contractile dysfunction (6.4±4.7% versus 12.6±9.6%; P=0.02). Over a median follow-up of 2.8 (1.2-5.7) years, LA contractile strain was the only significant predictor of AF in multivariable Cox regression (hazard ratio, 4.0 [95% CI, 1.04-15.2]). CONCLUSIONS: LMNA cardiomyopathy is associated with early intrinsic atrial myopathy reflected by high AF prevalence and reduced LA contractile strain, even in the absence of LV dysfunction and LA dilation. Whether LA strain can be used as a monitoring strategy to detect and mitigate AF complications requires validation.

Increased protein phosphatase 5 expression in inflammation-induced left ventricular dysfunction in rats.

BACKGROUND: Titin phosphorylation contributes to left ventricular (LV) diastolic dysfunction. The independent effects of inflammation on the molecular pathways that regulate titin phosphorylation are unclear. METHODS: We investigated the effects of collagen-induced inflammation and subsequent tumor necrosis factor-α (TNF-α) inhibition on mRNA expression of genes involved in regulating titin phosphorylation in 70 Sprague-Dawley rats. LV diastolic function was assessed with echocardiography. Circulating inflammatory markers were quantified by enzyme-linked immunosorbent assay and relative LV gene expression was assessed by Taqman® polymerase chain reaction. Differences in normally distributed variables between the groups were determined by two-way analysis of variance (ANOVA), followed by Tukey post-hoc tests. For non-normally distributed variables, group differences were determined by Kruskal-Wallis tests. RESULTS: Collagen inoculation increased LV relative mRNA expression of vascular cell adhesion molecule 1 (VCAM1), pentraxin 3 (PTX3), and inducible nitric oxide synthase (iNOS) compared to controls, indicating local microvascular inflammation. Collagen inoculation decreased soluble guanylate cyclase alpha-2 (sGCα2) and soluble guanylate cyclase beta-2 (sGCß2) expression, suggesting downregulation of nitric oxide-soluble guanylate cyclase-cyclic guanosine monophosphate (NO-sGC-cGMP) signaling. Inhibiting TNF-α prevented collagen-induced changes in VCAM1, iNOS, sGCα2 and sGCß2 expression. Collagen inoculation increased protein phosphatase 5 (PP5) expression. Like LV diastolic dysfunction, increased PP5 expression was not prevented by TNF-α inhibition. CONCLUSION: Inflammation-induced LV diastolic dysfunction may be mediated by a TNF-α-independent increase in PP5 expression and dephosphorylation of the N2-Bus stretch element of titin, rather than by TNF-α-induced downregulation of NO-sGC-cGMP pathway-dependent titin phosphorylation. The steady rise in number of patients with inflammation-induced diastolic dysfunction, coupled with low success rates of current therapies warrants a better understanding of the systemic signals and molecular pathways responsible for decreased titin phosphorylation in development of LV diastolic dysfunction. The therapeutic potential of inhibiting PP5 upregulation in LV diastolic dysfunction requires investigation.

Early Diagnosis of Abnormal Left Ventricular Systolic Functions of Rare Pathogenic Titin Mutation Gene Carriers in FHCM by Three-Dimensional Speckle Tracking Echocardiography Combined with Gene Detection.

Objective: This study aimed to explore the early diagnosis of abnormal left ventricular systolic function of rare pathogenic titin (TTN) mutation gene carriers in familial hypertrophic cardiomyopathy (FHCM) by three-dimensional speckle tracking echocardiography (3D-STE) combined with gene detection. Methods: Eighteen members of a Hui nationality family in Ningxia province of China were enrolled in this study in July 2019. The proband was tested with high-throughput sequencing of gene detection technology to detect the whole exome, and the mutation locus of pathogenic TTN gene was analyzed. According to the result, 16 subjects were divided into two groups: carrier group (n = 4) and noncarrier group (n = 12). Related indicators from 2DE were obtained, and myocardial strain indicators from 3D-STE were analyzed by postprocessing software of Tomtec. Strain indicators included global longitudinal strain (GLS), global circumference strain (GCS), global radial strain (GRS), regional longitudinal strain (RLS), regional circumference strain (RCS), and regional radial strain (RRS). All those indicators were compared between the two groups, and a receiver operating characteristic (ROC) curve was used for further analysis. Results: There were 4 subjects diagnosed as asymptomatic TTN gene carriers with the mutation locus of Val135643Ile. Compared with the noncarrier group, GLS and partial RLS were significantly reduced in the carrier group. The ROC curve shows that GLS has the largest AUC, and its sensitivity was better than LVPWD and specificity was better than IVSD and LVMI obtained from 2DE in the carrier group. Conclusions: There were 4 subjects diagnosed as asymptomatic TTN gene carriers with the mutation locus of Val135643Ile, and their GLS and partial RLS were significantly reduced; GLS had the better sensitivity and specificity than LVPWD, IVSD, and LVMI.

Relation of genetic polymorphisms in microRNAs with diastolic and systolic function in type 2 diabetes mellitus.

BACKGROUND AND AIMS: Type 2 diabetes mellitus (T2DM) has high risk of developing cardiac dysfunction, increasing of either cardiovascular death or hospitalization for heart failure. MicroRNAs (miRNA) affect cardiac function of T2DM. The aim of this study was to investigate the relationships between five miRNA single nucleotide polymorphisms (SNP) and diastolic and systolic function of T2DM. METHODS AND RESULTS: Three hundred untreated T2DM subjects were included. Each subject underwent SNP genotyping, conventional echocardiography, tissue doppler imaging, and speckle tracking imaging. The effects of miRNA SNPs on diastolic and systolic function were evaluated. The diastolic function of T2DM subjects with miR-133a-1-rs8089787 wild genotype or let-7f-rs10877887 variant genotype was lower than those with miR-133a-1-rs8089787 variant genotype or let-7f-rs10877887 wild genotype, manifesting as higher left atrial volume index, lower mean E', and higher E/E' (P < 0.05). There were no significant effects of miR-133a-2-rs13040413, let-7a-1-rs13293512 and miR-27a-rs895819 on the diastolic function of T2DM subjects (P > 0.05). These five miRNA SNPs had no effect on the systolic function of T2DM subjects (P > 0.05). CONCLUSIONS: MiRNA-133a-1-rs8089787 and let-7f-rs10877887 were associated with impaired cardiac diastolic function in T2DM. The findings may be a promising therapeutic targets for preventing diastolic dysfunction in T2DM.

Effects of idiopathic erythrocytosis on the left ventricular diastolic functions and the spectrum of genetic mutations: A case control study.

BACKGROUND: We have aimed at exposing left ventricular diastolic functions and the presence of known genetic mutations for familial erythrocytosis, in patients who exhibit idiopathic erythrocytosis. METHODS: Sixty-four patients with idiopathic erythrocytosis (mean age, 46.4 ± 2.7 years) and 30 age-matched healthy subjects were prospectively evaluated. The regions of interest of the erythropoietin receptor, hemoglobin beta-globin, von Hippel-Lindau, hypoxia-inducible factor 2 alpha, and Egl-9 family hypoxia-inducible factor genes were amplified by PCR. Left ventricular (LV) mass was measured by M-mode and 2-dimensional echocardiography. LV diastolic functions were assessed by conventional echocardiography and tissue Doppler imaging. RESULTS: As a result of genetic analyses, genetic mutations for familial erythrocytosis were detected in 5 patients. It has been observed in our study that the risk of cardiovascular disorders is higher in patients. Interventricular septum thickness, left atrial diameter, and some diastolic function parameters such as deceleration time and isovolumetric relaxation time have been found to be significantly higher in idiopathic erythrocytosis group than in the controls. CONCLUSION: This study has shown that LV diastolic functions were impaired in patients with idiopathic erythrocytosis. In this patient group with increased risk of cardiovascular disorders, the frequent genetic mutations have been detected in 5 patients only. Therefore, further clinical investigations are needed as novel genetic mutations may be discovered in patients with idiopathic erythrocytosis because of cardiovascular risk.

The interleukin-1 receptor type I promotes the development of aging-associated cardiomyopathy in mice.

BACKGROUND: Aging is associated with metabolic and structural changes causing heart failure with preserved ejection fraction (HFpEF). Interleukin-1 (IL-1) is a pro-inflammatory cytokine involved in aging-related inflammation. OBJECTIVE: We sought to determine whether IL-1 mediates aging-related changes in the heart, as seen in HFpEF. METHODS: We studied age-matched young (4-month-old), middle-aged (14-month-old), and old (23-month-old) wild-type (WT) C57BL/6J and IL-1 receptor type I deficient (IL1RI-KO) male mice. Echocardiography was used to evaluate left ventricular (LV) dimensions and systolic/diastolic function, and a pressure transducer was used to measure the LV end-diastolic pressure. Picrosirius red stain was used to assess for myocardial interstitial fibrosis (MIF) at pathology. RESULTS: WT and IL-1RIKO mice showed a normal cardiac phenotype at young age, without any differences between the two groups. With aging, the WT mice developed LV concentric hypertrophy (as measured by a significant increase in LV mass [+42%, P < 0.01] and relative wall thickness [+34%, P < 0.01]), whereas the aging IL-1RI-KO mice did not. With aging, the WT mice also developed diastolic dysfunction (as measured by a significant increase in isovolumetric relaxation time [+148%, P < 0.01] and a significantly higher LV end-diastolic pressure [+174%, P < 0.01]), whereas the aging IL1RI-KO did not. Aged WT mice showed a significant increase in MIF (+124%, P < 0.01) at cardiac pathology, whereas the aging IL-1RI-KO did not. CONCLUSIONS: Genetically-modified mice lacking the IL-1RI receptor, not responsive to IL-1, are protected from aging-related LV hypertrophy, fibrosis, and diastolic dysfunction. These data support a central role of IL-1 in the pathophysiology of aging-related HFpEF.

Polymorphic Variants in the GRK5 Gene Promoter Are Associated With Diastolic Dysfunction in Coronary Artery Bypass Graft Surgery Patients.

BACKGROUND: The G-protein-coupled receptor kinase 5 (GRK5) is a mediator of cardiovascular homeostasis and participates in inflammation and cardiac fibrosis, both being involved in the development of diastolic dysfunction (DD). While mechanisms of transcriptional regulation of the GRK5 promoter are unclear, we tested the hypotheses, that (1) GRK5 expression varies depending on functional single nucleotide polymorphisms (SNPs) in the GRK5 promoter and (2) this is associated with DD in patients undergoing coronary artery bypass graft (CABG) surgery. METHODS: We amplified and sequenced the GRK5 promoter followed by cloning, reporter assays, and electrophoretic mobility shift assays (EMSA). GRK5 messenger ribonucleic acid (mRNA) expression was determined in right atrial tissue sampled from 50 patients undergoing CABG surgery. In another prospective study, GRK5 genotypes were associated with determinants of diastolic function using transesophageal echocardiography in 255 patients with CABG with normal systolic left ventricular (LV) function. Specifically, we measured ejection fraction (EF), transmitral Doppler early filling velocity (E), tissue Doppler early diastolic lateral mitral annular velocity (E' lateral), and calculated E/E', E' norm and the difference of E' lateral and E' norm to account for age-related changes in diastolic function. RESULTS: We identified 6 SNPs creating 3 novel haplotypes with the greatest promoter activation in haplotype tagging (ht) SNP T(-678)C T-allele constructs (P < .001). EMSAs showed allele-specific transcription factor binding proving functional activity. GRK5 mRNA expression was greatest in TT genotypes (TT: 131 fg/µg [95% CI, 108-154]; CT: 109 [95% confidence interval {CI}, 93-124]; CC: 83 [95% CI, 54-112]; P = .012). Moreover, GRK5 genotypes were significantly associated with determinants of diastolic function. Grading of DD revealed more grade 3 patients in TT compared to CT and CC genotypes (58% vs 38% vs 4%; P = .023). E´ lateral was lowest in TT genotypes (P = .007) and corresponding E/E' measurements showed 1.27-fold increased values in TT versus CC genotypes (P = .01), respectively. While E' norm values were not different between genotypes (P = .182), the difference between E' lateral and E' norm was significantly higher in TT genotypes compared to CC and CT genotypes (-1.2 [interquartile range {IQR}, 2.7], -0.5 [IQR, 3.4], and -0.4 [IQR, 4.2; P = .035], respectively). CONCLUSIONS: A functional GRK5 SNP results in allele-dependent differences in GRK5 promoter activity and mRNA expression. This is associated with altered echocardiographic determinants of diastolic function. Thus, SNPs in the GRK5 promoter are associated with altered perioperative diastolic cardiac function. In the future, preoperative testing for these and other SNPs might allow to initiate more specific diagnostic and perioperative pathways to benefit patients at risk.